Spontaneously occurring autoantibodies, such as antinuclear antibodies, which are found in human diseases such as systemic lupus erythematosus have been used as reagents to identify nuclear antigens which are reactive with these antibodies and in turn using these nuclear antigens to characterize the biochemical specificities of different types of antinuclear antibodies. In this connection, anti-DNA antibodies, anti-deoxyribonucleoprotein antibodies and anti-nucleolar antibodies were used to identify and to locate the respective subcellular macromolecules at different phases of the cell cycle. These studies demonstrated that native DNA and deoxyribonucleoprotein were located primarily in the nuclear periphery during the G1 phase of the cell cycle. Cytoplasmic DNA was present in the plasma membrane especially during metaphase. The importance of deoxyribonucleoprotein and antibodies to deoxyribonucleoprotein as potential reactants in antigen-antibody reactions in systemic lupus erythematosus and in rheumatoid arthritis was demonstrated in recent studies. A nucleolar macromolecule, a 4-6S RNA which is nucleolus specific, was demonstrated to be an antigen which was reactive with antinucleolar antibodies present in the sera of patients with progressive systemic sclerosis and Raynaud's phenomenon. The 4-6S RNA could be isolated from the nucleoli of tissues such as rat liver and calf thymus and could be used to absorb the nucleolar antibodies present in such sera.